(a) Field of the Invention
The invention relates to a new primer pair consisting of R14B/264 primer and a primer designed from Sextolet 900 marker and to a highly informative DNA marker obtained from amplification of genomic DNA with the primer pair.
(b) Description of Prior Art
The science of molecular genetics provides researchers with the tremendous opportunities to study phenomena underlying genetic diseases and individualization. Both stability and variability of biological characteristics of all living organisms are assured by transmission of the hereditary material (DNA) between generations. However, the evolution process brings about cumulative differences between individuals at various levels. Sufficient data obtained from the study of polymorphic parts of the DNA (genetic markers) in human have helped map part of the human genome, and identify genes responsible for particular diseases. In addition, genetic markers are used to differentiate between individuals at the molecular level.
The basis of DNA polymorphism derives from differences in DNA sequences inherited from each parental chromosome. DNA polymorphism could also arise from a mutation in one defined single nucleotide, an insertion or a deletion of a sequence, or from variations in the length of a stretch of short tandem repeats (STR). Polymorphic DNA markers provide information regarding the segregation pattern of parental chromosomes during the mating process and hence disclose a person's genetic identity. The informativity of a specific marker is measured by its heterozigosity (H), its polymorphic information content (PIC) and the allelic frequency of each allele. The total informativity of a marker system (where several markers may be involved) is a function of the informativity extracted from the individual markers. The higher informativity of individual markers, the more informative a marker system is.
Several methods have been developed to evaluate DNA polymorphism. Restriction fragment length polymorphism (RFLP) based on Southern Blot technique is widely used to identify DNA polymorphism related to genetic diseases, genome mapping, evolution and forensic studies. Microsatellite DNA markers revealed by this method are highly informative. However, this method requires considerable amounts of DNA and large sample sizes. In addition, relatively long analysis times are needed to obtain results and translate into high cost and increases in turn around time.
Using the polymerase chain reaction (PCR), DNA present in minute amounts of biological material can be amplified into as many copies as needed. Amplified fragments are easy to be analyzed. The application of PCR to DNA marker analysis proves to be fast, reliable and cost effective. Most markers which have thus far been analyzed by PCR are of STR polymorphism. These markers consist of more than two alleles with the exceptions of some highly informative ones where more than 35 alleles (Kimpton C., et al. Forensic Science International 71(2):137-52, 1995) and a heterozigosity less than 0.93 have been described. No marker with more than 40 alleles and heterozigosity greater than 0.96 has been described so far.
Marker Q900 described by Tang et al. (Tang J. Q., et al., Mammalian Genome 6:345-349, 1995) reportedly consisted of 23 alleles and had a heterozigosity of 0.95.
U.S. Pat. No. 5,576,180 in the name of Melancon et al. discloses a DNA amplification primer pair for the simultaneous amplification of multiple highly polymorphic genomic loci. The primer pair consists of R14B/264 and Q560mak. This primer pair allows for the identification of three (3) markers including the marker Q900. However, only 23 alleles with a heterozigosity of 0.95 have been observed using the marker Q900.
It would be highly desirable to be provided with a primer pair allowing for the amplification of a marker having a high discrimination power and heterozigosity.
It would be highly desirable to be provided with a reliable method for DNA fingerprinting identification of genetically related or unrelated individuals.